Physiological And Proteomics Analyses Of Chloroplasts From Spinacia Oleracea L.in Response To H₂O₂

Oxidative stress caused by various environmental stresses is one of the main factors affecting plant growth.H₂O₂,a reactive oxygen species（ROS）,has certain damage to normal physiological metabolism of plant cells and is also an important regulatory molecule involved in signal transduction.The investigation of the dynamic changes of the protein redox status during the H₂O₂ response is important for dissecting the ROS-regulated signals and various metabolic pathways in plants.Spinach（Spinacia oleracea L.）,a common green leafy vegetables belong to genus Spinacia of Amaranth family,is a good model for studying plant photosynthesis mechanism.Importantly,spinach is widely distributed and one of the key vegetables in China.Studying the stress-responsive molecular mechanism in spinach is vital for molecular maker-assisted breeding.Using physiological and redox proteomics approaches,we analyzed the signal and metabolic characteristics of chloroplasts in spinach leaves in response to H₂O₂（0 mM,5 mM,and 10 mM for 20 min）stresses.We found 61 differential abundant proteins and 25 differential redox level proteins in response to H₂O₂,using isobaric tags for relative and absolute quantification（iTRAQ）and thiol-based specific tandem mass tag（iodoTMT）.These proteins were mainly invovled in signal transduction,photosynthesis,carbon and energy metabolism,stress defense,and protein turnover.The abundances and redox status of these proteins preliminarily revealed various oxidative stress-responsive signal and metabolism mechanism in chloroplasts,including:（1）reducing light energy capture and absorption,inhibiting photosynthetic electron transport and Calvin cycle,and reducing photosynthetic rate;（2）regulating the redox status of photosynthetic proteins,thereby affecting protein conformation and function to cope with H₂O₂ stress;（3）initiating GSH-GSSG pathway in glutathione in ascorbate-glutathione（ASA-GSH）cycle to scavenge ROS;（4）enhancing glycolysis and the Krebs cycle pathway for energy supply;（5）promoting some basal metabolic pathways;（6）modify the patterns of protein translation and processing to deal with oxidative stress-induced protein damage.These results provide valuable information for further studies on the oxidative stress-responsive signal and metabolism in spinach chloroplasts.